Bone-derived soluble factors and laminin-511 cooperate to promote migration, invasion and survival of bone-metastatic breast tumor cells

Abstract

Tumor intrinsic and extrinsic factors are thought to contribute to bone metastasis but little is known about how they cooperate to promote breast cancer spread to bone. We used the bone-metastatic 4T1BM2 mammary carcinoma model to investigate the cooperative interactions between tumor LM-511 and bone-derived soluble factors in vitro. We show that bone conditioned medium cooperates with LM-511 to enhance 4T1BM2 cell migration and invasion and is sufficient alone to promote survival in the absence of serum. These responses were associated with increased secretion of MMP-9 and activation of ERK and AKT signaling pathways and were partially blocked by pharmacological inhibitors of MMP-9, AKT-1/2 or MEK. Importantly, pre-treatment of 4T1BM2 cells with an AKT-1/2 inhibitor significantly reduced experimental metastasis to bone in vivo. Promotion of survival and invasive responses by bone-derived soluble factors and tumor-derived LM-511 are likely to contribute to the metastatic spread of breast tumors to bone.

Identification of brain metastasis genes and therapeutic evaluation of histone deacetylase inhibitors in a clinically relevant model of breast cancer brain metastasis

“…Western blotting was performed using standard methodology as described previously 694 (Denoyer et al, 2014). Briefly, 40 µg of proteins were separated on a 15% SDS-PAGE gel 695 and transferred to a PVDF membrane.…”

“…Statistical 731 difference between groups was analysed using a Mann-Whitney nonparametric test when 732 comparing two groups or a one-way ANOVA, Bonferroni post-test when comparing multiple 733 groups; p < 0.05 was considered significant. 734 735Proliferation assay and determination of inhibitory concentration (IC 50 ) 736Cell proliferation was measured using a sulforhodamine B (SRB) colorimetric assay as 737 described previously(Denoyer et al, 2014). Proliferation was measured over 5 days in SFM 738 supplemented with 50% v/v BCM or 1% FBS or both.…”

Neoadjuvant neratinib promotes ferroptosis and inhibits brain metastasis in a novel syngeneic model of spontaneous HER2+ve breast cancer metastasis

“…Cell proliferation was measured using a sulforhodamine B (SRB) colorimetric assay as described previously with minor modifications [34,37]. Briefly, TBCP-1 (1 × 10 3 ) or SKBR3 (2 × 10 3 ) cells were seeded in triplicate wells of a 96-well plate in 100 μl of serum-containing medium and allowed to adhere at 37°C for 6 h. Inhibitors were added in 100 μl of the same medium and proliferation measured over 5 days.…”

“…Expression of ERα, PR and HER2 in sub-confluent cultures of TBCP-1 cells was detected by standard immunoblotting [37]. Primary antibodies against ERα (Santa Cruz sc-542, 1 μg/ml), PR (Santa Cruz sc-538, 1 μg/ml) or HER2 (Abcam ab2428, 1 μg/ml) and appropriate horseradish peroxidase (HRP)-conjugated secondary antibodies were used to detect the respective proteins.…”

“…Experimental metastasis assays were done as described previously with minor modifications [34,37]. For initial characterisation of metastatic spread, TBCP-1 cells (5 × 10 5 ) were injected into the left cardiac ventricle of 6-8week-old female BALB/C mice.…”